JoVE Visualize What is visualize?
Stop Reading. Start Watching.
Advanced Search
Stop Reading. Start Watching.
Regular Search
Find video protocols related to scientific articles indexed in Pubmed.
Promoter Activation by CII, a Potent Transcriptional Activator from Bacteriophage 186.
J. Biol. Chem.
PUBLISHED: 10-06-2014
Show Abstract
Hide Abstract
The lysogeny promoting protein CII from bacteriophage 186 is a potent transcriptional activator, capable of mediating at least a 400-fold increase in transcription over basal activity. Despite being functionally similar to its counterpart in phage ?, it shows no homology at the level of protein sequence and does not belong to any known family of transcriptional activators. It also has the unusual property of binding DNA half-sites that are separated by 20 base pairs, center to center. Here we investigate the structural and functional properties of CII using a combination of genetics, in vitro assays, and mutational analysis. We find that 186 CII possesses two functional domains, with an independent activation epitope in each. 186 CII owes its potent activity to activation mechanisms that are dependent on both the ?(70) and ? C-terminal domain (?CTD) components of RNA polymerase, contacting different functional domains. We also present evidence that like ? CII, 186 CII is proteolytically degraded in vivo, but unlike ? CII, 186 CII proteolysis results in a specific, transcriptionally inactive, degradation product with altered self-association properties.
Related JoVE Video
Quantitation of interactions between two DNA loops demonstrates loop domain insulation in E. coli cells.
Proc. Natl. Acad. Sci. U.S.A.
PUBLISHED: 10-06-2014
Show Abstract
Hide Abstract
Eukaryotic gene regulation involves complex patterns of long-range DNA-looping interactions between enhancers and promoters, but how these specific interactions are achieved is poorly understood. Models that posit other DNA loops-that aid or inhibit enhancer-promoter contact-are difficult to test or quantitate rigorously in eukaryotic cells. Here, we use the well-characterized DNA-looping proteins Lac repressor and phage ? CI to measure interactions between pairs of long DNA loops in E. coli cells in the three possible topological arrangements. We find that side-by-side loops do not affect each other. Nested loops assist each other's formation consistent with their distance-shortening effect. In contrast, alternating loops, where one looping element is placed within the other DNA loop, inhibit each other's formation, thus providing clear support for the loop domain model for insulation. Modeling shows that combining loop assistance and loop interference can provide strong specificity in long-range interactions.
Related JoVE Video
Road rules for traffic on DNA-systematic analysis of transcriptional roadblocking in vivo.
Nucleic Acids Res.
PUBLISHED: 07-17-2014
Show Abstract
Hide Abstract
Genomic DNA is bound by many proteins that could potentially impede elongation of RNA polymerase (RNAP), but the factors determining the magnitude of transcriptional roadblocking in vivo are poorly understood. Through systematic experiments and modeling, we analyse how roadblocking by the lac repressor (LacI) in Escherichia coli cells is controlled by promoter firing rate, the concentration and affinity of the roadblocker protein, the transcription-coupled repair protein Mfd, and promoter-roadblock spacing. Increased readthrough of the roadblock at higher RNAP fluxes requires active dislodgement of LacI by multiple RNAPs. However, this RNAP cooperation effect occurs only for strong promoters because roadblock-paused RNAP is quickly terminated by Mfd. The results are most consistent with a single RNAP also sometimes dislodging LacI, though we cannot exclude the possibility that a single RNAP reads through by waiting for spontaneous LacI dissociation. Reducing the occupancy of the roadblock site by increasing the LacI off-rate (weakening the operator) increased dislodgement strongly, giving a stronger effect on readthrough than decreasing the LacI on-rate (decreasing LacI concentration). Thus, protein binding kinetics can be tuned to maintain site occupation while reducing detrimental roadblocking.
Related JoVE Video
Long-term impact of deficit irrigation on the physical quality of berries in 'Crimson Seedless' table grapes.
J. Sci. Food Agric.
PUBLISHED: 06-30-2014
Show Abstract
Hide Abstract
In table grapes, berry firmness influences consumer acceptance so it is important to avoid berry shattering and dehydration during their postharvest life. Since studies of irrigation effects on table grape quality are comparatively rare, sensory evaluation aimed to identify high-quality berries obtained under different deficit irrigation (DI) treatments. A three-year study examined the effects of DI strategies on some physical quality attributes at harvest, after 28 days of cold storage at 0?°C and after an additional shelf-life period of 3 d at 15?°C. Control vines were irrigated to ensure non-limiting water conditions (110% of crop evapotranspiration, ETc), while both regulated deficit irrigation treatment (RDI) and partial rootzone drying (PRD) treatments applied 35% less water post-veraison. The null irrigation treatment (NI) only received natural precipitation (72% less water than control vines).
Related JoVE Video
Cytokinin producing bacteria stimulate amino acid deposition by wheat roots.
Plant Physiol. Biochem.
PUBLISHED: 03-06-2014
Show Abstract
Hide Abstract
Phytohormone production is one mechanism by which rhizobacteria can stimulate plant growth, but it is not clear whether the bacteria gain from this mechanism. The hypothesis that microbial-derived cytokinin phytohormones stimulate root exudation of amino acids was tested. The rhizosphere of wheat plants was drenched with the synthetic cytokinin trans-zeatin or inoculated with Bacillus subtilis IB-22 (which produces zeatin type cytokinins) or B. subtilis IB-21 (which failed to accumulate cytokinins). Growing plants in a split root system allowed spatial separation of zeatin application or rhizobacterial inoculation to one compartment and analyses of amino acid release from roots (rhizodeposition) into the other compartment (without either microbial inoculation or treatment with exogenous hormone). Supplying B. subtilis IB-22 or zeatin to either the whole root system or half of the roots increased concentrations of amino acids in the soil solution although the magnitude of the increase was greater when whole roots were treated. There was some similarity in amino acid concentrations induced by either bacterial or zeatin treatment. Thus B. subtilis IB-22 increased amino acid rhizodeposition, likely due to its ability to produce cytokinins. Furthermore, B. subtilis strain IB-21, which failed to accumulate cytokinins in culture media, did not significantly affect amino acid concentrations in the wheat rhizosphere. The ability of rhizobacteria to produce cytokinins and thereby stimulate rhizodeposition may be important in enhancing rhizobacterial colonization of the rhizoplane.
Related JoVE Video
Physiological and gene expression responses of sunflower (Helianthus annuus L.) plants differ according to irrigation placement.
Plant Sci.
PUBLISHED: 03-03-2014
Show Abstract
Hide Abstract
To investigate effects of soil moisture heterogeneity on plant physiology and gene expression in roots and leaves, three treatments were implemented in sunflower plants growing with roots split between two compartments: a control (C) treatment supplying 100% of plant evapotranspiration, and two treatments receiving 50% of plant evapotranspiration, either evenly distributed to both compartments (deficit irrigation - DI) or unevenly distributed to ensure distinct wet and dry compartments (partial rootzone drying - PRD). Plants receiving the same amount of water responded differently under the two irrigation systems. After 3 days, evapotranspiration was similar in C and DI, but 20% less in PRD, concomitant with decreased leaf water potential (?leaf) and increased leaf xylem ABA concentration. Six water-stress responsive genes were highly induced in roots growing in the drying soil compartment of PRD plants, and their expression was best correlated with local soil water content. On the other hand, foliar gene expression differed significantly from that of the root and correlated better with xylem ABA concentration and ?leaf. While the PRD irrigation strategy triggered stronger physiological and molecular responses, suggesting a more intense and systemic stress reaction due to local dehydration of the dry compartment of PRD plants, the DI strategy resulted in similar water savings without strongly inducing these responses. Correlating physiological and molecular responses in PRD/DI plants may provide insights into the severity and location of water deficits and may enable a better understanding of long-distance signalling mechanisms.
Related JoVE Video
Quantitation of the DNA tethering effect in long-range DNA looping in vivo and in vitro using the Lac and ? repressors.
Proc. Natl. Acad. Sci. U.S.A.
PUBLISHED: 12-16-2013
Show Abstract
Hide Abstract
Efficient and specific interactions between proteins bound to the same DNA molecule can be dependent on the length of the DNA tether that connects them. Measurement of the strength of this DNA tethering effect has been largely confined to short separations between sites, and it is not clear how it contributes to long-range DNA looping interactions, such as occur over separations of tens to hundreds of kilobase pairs in vivo. Here, gene regulation experiments using the LacI and ? CI repressors, combined with mathematical modeling, were used to quantitate DNA tethering inside Escherichia coli cells over the 250- to 10,000-bp range. Although LacI and CI loop DNA in distinct ways, measurements of the tethering effect were very similar for both proteins. Tethering strength decreased with increasing separation, but even at 5- to 10-kb distances, was able to increase contact probability 10- to 20-fold and drive efficient looping. Tethering in vitro with the Lac repressor was measured for the same 600-to 3,200-bp DNAs using tethered particle motion, a single molecule technique, and was 5- to 45-fold weaker than in vivo over this range. Thus, the enhancement of looping seen previously in vivo at separations below 500 bp extends to large separations, underlining the need to understand how in vivo factors aid DNA looping. Our analysis also suggests how efficient and specific looping could be achieved over very long DNA separations, such as what occurs between enhancers and promoters in eukaryotic cells.
Related JoVE Video
Structural-functional dissection and characterization of yield-contributing traits originating from a group 7 chromosome of the wheatgrass species Thinopyrum ponticum after transfer into durum wheat.
J. Exp. Bot.
PUBLISHED: 12-06-2013
Show Abstract
Hide Abstract
For the first time, using chromosome engineering of durum wheat, the underlying genetic determinants of a yield-improving segment from Thinopyrum ponticum (7AgL) were dissected. Three durum wheat-Th. ponticum near-isogenic recombinant lines (NIRLs), with distal portions of their 7AL arm (fractional lengths 0.77, 0.72, and 0.60) replaced by alien chromatin, were field-tested for two seasons under rainfed conditions. Yield traits and other agronomic characteristics of the main shoot and whole plant were measured. Loci for seed number per ear and per spikelet were detected in the proximal 7AgL segment (0.60-0.72). Loci determining considerable increases of flag leaf width and area, productive tiller number per plant, biomass per plant, and grain yield per plant were located in the distally adjacent 0.72-0.77 7AgL segment, while in the most distal portion (0.77-1.00) genetic effects on spikelet number per ear were identified. Contrary to previous reports, trials with the bread wheat T4 translocation line, carrying on 7DL a sizeable 7AgL segment of which those present in the durum wheat-Th. ponticum NIRLs represent fractions, gave no yield advantage. The hypothesis that ABA might be a factor contributing to the 7AgL effects was tested by analysing endogenous ABA contents of the NIRLs and their responses to exogenous ABA application. The 7AgL yield-related loci were shown to be ABA-independent. This study highlights the value of wheat-alien recombinant lines for dissecting the genetic and physiological basis of complex traits present in wild germplasm, and provides a basis for their targeted exploitation in wheat breeding.
Related JoVE Video
Collaboration between CpG sites is needed for stable somatic inheritance of DNA methylation states.
Nucleic Acids Res.
PUBLISHED: 11-27-2013
Show Abstract
Hide Abstract
Inheritance of 5-methyl cytosine modification of CpG (CG/CG) DNA sequences is needed to maintain early developmental decisions in vertebrates. The standard inheritance model treats CpGs as independent, with methylated CpGs maintained by efficient methylation of hemimethylated CpGs produced after DNA replication, and unmethylated CpGs maintained by an absence of de novo methylation. By stochastic simulations of CpG islands over multiple cell cycles and systematic sampling of reaction parameters, we show that the standard model is inconsistent with many experimental observations. In contrast, dynamic collaboration between CpGs can provide strong error-tolerant somatic inheritance of both hypermethylated and hypomethylated states of a cluster of CpGs, reproducing observed stable bimodal methylation patterns. Known recruitment of methylating enzymes by methylated CpGs could provide the necessary collaboration, but we predict that recruitment of demethylating enzymes by unmethylated CpGs strengthens inheritance and allows CpG islands to remain hypomethylated within a sea of hypermethylation.
Related JoVE Video
Abscisic acid metabolizing rhizobacteria decrease ABA concentrations in planta and alter plant growth.
Plant Physiol. Biochem.
PUBLISHED: 09-22-2013
Show Abstract
Hide Abstract
Although endogenous phytohormones such as abscisic acid (ABA) regulate root growth, and many rhizobacteria can modulate root phytohormone status, hitherto there have been no reports of rhizobacteria mediating root ABA concentrations and growth by metabolising ABA. Using a selective ABA-supplemented medium, two bacterial strains were isolated from the rhizosphere of rice (Oryza sativa) seedlings grown in sod-podzolic soil and assigned to Rhodococcus sp. P1Y and Novosphingobium sp. P6W using partial 16S rRNA gene sequencing and phenotypic patterns by the GEN III MicroPlate test. Although strain P6W had more rapid growth in ABA-supplemented media than strain P1Y, both could utilize ABA as a sole carbon source in batch culture. When rice seeds were germinated on filter paper in association with bacteria, root ABA concentration was not affected, but shoot ABA concentration of inoculated plants decreased by 14% (strain P6W) and 22% (strain P1Y). When tomato (Solanum lycopersicum) genotypes differing in ABA biosynthesis (ABA deficient mutants flacca - flc, and notabilis - not and the wild-type cv. Ailsa Craig, WT) were grown in gnotobiotic cultures on nutrient solution agar, rhizobacterial inoculation decreased root and/or leaf ABA concentrations, depending on plant and bacteria genotypes. Strain P6W inhibited primary root elongation of all genotypes, but increased leaf biomass of WT plants. In WT plants treated with silver ions that inhibit ethylene perception, both ABA-metabolising strains significantly decreased root ABA concentration, and strain P6W decreased leaf ABA concentration. Since these changes in ABA status also occurred in plants that were not treated with silver, it suggests that ethylene was probably not involved in regulating bacteria-mediated changes in ABA concentration. Correlations between plant growth and ABA concentrations in planta suggest that ABA-metabolising rhizobacteria may stimulate growth via an ABA-dependent mechanism.
Related JoVE Video
How do roots elongate in a structured soil?
J. Exp. Bot.
PUBLISHED: 09-16-2013
Show Abstract
Hide Abstract
In this review, we examine how roots penetrate a structured soil. We first examine the relationship between soil water status and its mechanical strength, as well as the ability of the soil to supply water to the root. We identify these as critical soil factors, because it is primarily in drying soil that mechanical constraints limit root elongation. Water supply to the root is important because root water status affects growth pressures and root stiffness. To simplify the bewildering complexity of soil-root interactions, the discussion is focused around the special cases of root elongation in soil with pores much smaller than the root diameter and the penetration of roots at interfaces within the soil. While it is often assumed that the former case is well understood, many unanswered questions remain. While low soil-root friction is often viewed as a trait conferring better penetration of strong soils, it may also increase the axial pressure on the root tip and in so doing reduce the rate of cell division and/or expansion. The precise trade-off between various root traits involved in root elongation in homogeneous soil remains to be determined. There is consensus that the most important factors determining root penetration at an interface are the angle at which the root attempts to penetrate the soil, root stiffness, and the strength of the soil to be penetrated. The effect of growth angle on root penetration implicates gravitropic responses in improved root penetration ability. Although there is no work that has explored the effect of the strength of the gravitropic responses on penetration of hard layers, we attempt to outline possible interactions. Impacts of soil drying and strength on phytohormone concentrations in roots, and consequent root-to-shoot signalling, are also considered.
Related JoVE Video
Bacteriophage lambda repressor mediates the formation of a complex enhancer-like structure.
Transcription
PUBLISHED: 08-31-2013
Show Abstract
Hide Abstract
For the past 40 years, bacteriophage lambda has been crucial in revealing fundamental principles underlying control of transcription by elements positioned close to promoters. With the discovery that lambda CI repressors bound to distant sites can interact efficiently, lambda also provides a model for long range gene regulation, including the action of enhancer elements.
Related JoVE Video
Isoprene emission protects photosynthesis but reduces plant productivity during drought in transgenic tobacco (Nicotiana tabacum) plants.
New Phytol.
PUBLISHED: 05-23-2013
Show Abstract
Hide Abstract
Isoprene protects the photosynthetic apparatus of isoprene-emitting plants from oxidative stress. The role of isoprene in the response of plants to drought is less clear. Water was withheld from transgenic isoprene-emitting and non-emitting tobacco (Nicotiana tabacum) plants, to examine: the response of isoprene emission to plant water deficit; a possible relationship between concentrations of the drought-induced phytohormone abscisic acid (ABA) and isoprene; and whether isoprene affected foliar reactive oxygen species (ROS) and lipid peroxidation levels. Isoprene emission did not affect whole-plant water use, foliar ABA concentration or leaf water potential under water deficit. Compared with well-watered controls, droughted non-emitting plants significantly increased ROS content (31-46%) and lipid peroxidation (30-47%), concomitant with decreased operating and maximum efficiencies of photosystem II photochemistry and lower leaf and whole-plant water use efficiency (WUE). Droughted isoprene-emitting plants showed no increase in ROS content or lipid peroxidation relative to well-watered controls, despite isoprene emission decreasing before leaf wilting. Although isoprene emission protected the photosynthetic apparatus and enhanced leaf and whole-plant WUE, non-emitting plants had 8-24% more biomass under drought, implying that isoprene emission incurred a yield penalty.
Related JoVE Video
One-step cloning and chromosomal integration of DNA.
ACS Synth Biol
PUBLISHED: 05-20-2013
Show Abstract
Hide Abstract
We describe "clonetegration", a method for integrating DNA into prokaryotic chromosomes that approaches the simplicity of cloning DNA within extrachromosomal vectors. Compared to existing techniques, clonetegration drastically decreases the time and effort needed for integration of single or multiple DNA fragments. Additionally, clonetegration facilitates cloning and expression of genetic elements that are impossible to propagate within typical multicopy plasmids.
Related JoVE Video
Single molecule analysis of DNA wrapping and looping by a circular 14mer wheel of the bacteriophage 186 CI repressor.
Nucleic Acids Res.
PUBLISHED: 04-24-2013
Show Abstract
Hide Abstract
The lytic-lysogenic decision in bacteriophage 186 is governed by the 186 CI repressor protein in a unique way. The 186 CI is proposed to form a wheel-like oligomer that can mediate either wrapped or looped nucleoprotein complexes to provide the cooperative and competitive interactions needed for regulation. Although consistent with structural, biochemical and gene expression data, many aspects of this model are based on inference. Here, we use atomic force microscopy (AFM) to reveal the various predicted wrapped and looped species, and new ones, for CI regulation of lytic and lysogenic transcription. Automated AFM analysis showed CI particles of the predicted dimensions on the DNA, with CI multimerization favoured by DNA binding. Measurement of the length of the wrapped DNA segments indicated that CI may move on the DNA, wrapping or releasing DNA on either side of the wheel. Tethered particle motion experiments were consistent with wrapping and looping of DNA by CI in solution, where in contrast to ? repressor, the looped species were exceptionally stable. The CI regulatory system provides an intriguing comparison with that of nucleosomes, which share the ability to wrap and release similar sized segments of DNA.
Related JoVE Video
The rhizobacterium Variovorax paradoxus 5C-2, containing ACC deaminase, promotes growth and development of Arabidopsis thaliana via an ethylene-dependent pathway.
J. Exp. Bot.
PUBLISHED: 02-11-2013
Show Abstract
Hide Abstract
Many plant-growth-promoting rhizobacteria (PGPR) associated with plant roots contain the enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase and can metabolize ACC, the immediate precursor of the plant hormone ethylene, thereby decreasing plant ethylene production and increasing plant growth. However, relatively few studies have explicitly linked ethylene emission and/or action to growth promotion in these plant-microbe interactions. This study examined effects of the PGPR Variovorax paradoxus 5C-2 containing ACC deaminase on the growth and development of Arabidopsis thaliana using wild-type (WT) plants and several ethylene-related mutants (etr1-1, ein2-1, and eto1-1). Soil inoculation with V. paradoxus 5C-2 promoted growth (leaf area and shoot biomass) of WT plants and the ethylene-overproducing mutant eto1-1, and also enhanced floral initiation of WT plants by 2.5 days. However, these effects were not seen in ethylene-insensitive mutants (etr1-1 and ein2-1) even though bacterial colonization of the root system was similar. Furthermore, V. paradoxus 5C-2 decreased ACC concentrations of rosette leaves of WT plants by 59% and foliar ethylene emission of both WT plants and eto1-1 mutants by 42 and 37%, respectively. Taken together, these results demonstrate that a fully functional ethylene signal transduction pathway is required for V. paradoxus 5C-2 to stimulate leaf growth and flowering of A. thaliana.
Related JoVE Video
Enhancer-like long-range transcriptional activation by ? CI-mediated DNA looping.
Proc. Natl. Acad. Sci. U.S.A.
PUBLISHED: 02-04-2013
Show Abstract
Hide Abstract
How distant enhancer elements regulate the assembly of a transcription complex at a promoter remains poorly understood. Here, we use long-range gene regulation by the bacteriophage ? CI protein as a powerful system to examine this process in vivo. A 2.3-kb DNA loop, formed by CI bridging its binding sites at OR and OL, is known already to enhance repression at the lysogenic promoter PRM, located at OR. Here, we show that CI looping also activates PRM by allowing the C-terminal domain of the ? subunit of the RNA polymerase bound at PRM to contact a DNA site adjacent to the distal CI sites at OL. Our results establish OL as a multifaceted enhancer element, able to activate transcription from long distances independently of orientation and position. We develop a physicochemical model of our in vivo data and use it to show that the observed activation is consistent with a simple recruitment mechanism, where the ?-C-terminal domain to DNA contact need only provide ?2.7 kcal/mol of additional binding energy for RNA polymerase. Structural modeling of this complete enhancer-promoter complex reveals how the contact is achieved and regulated, and suggests that distal enhancer elements, once appropriately positioned at the promoter, can function in essentially the same way as proximal promoter elements.
Related JoVE Video
Long-distance abscisic acid signalling under different vertical soil moisture gradients depends on bulk root water potential and average soil water content in the root zone.
Plant Cell Environ.
PUBLISHED: 01-25-2013
Show Abstract
Hide Abstract
To determine how root-to-shoot abscisic acid (ABA) signalling is regulated by vertical soil moisture gradients, root ABA concentration ([ABA](root)), the fraction of root water uptake from, and root water potential of different parts of the root zone, along with bulk root water potential, were measured to test various predictive models of root xylem ABA concentration [RX-ABA](sap). Beans (Phaseolus vulgaris?L. cv. Nassau) were grown in soil columns and received different irrigation treatments (top and basal watering, and withholding water for varying lengths of time) to induce different vertical soil moisture gradients. Root water uptake was measured at four positions within the column by continuously recording volumetric soil water content (?v). Average ?v was inversely related to bulk root water potential (?(root)). In turn, ?(root) was correlated with both average [ABA](root) and [RX-ABA](sap). Despite large gradients in ?v, [ABA](root) and root water potential was homogenous within the root zone. Consequently, unlike some split-root studies, root water uptake fraction from layers with different soil moisture did not influence xylem sap (ABA). This suggests two different patterns of ABA signalling, depending on how soil moisture heterogeneity is distributed within the root zone, which might have implications for implementing water-saving irrigation techniques.
Related JoVE Video
Ethylene limits abscisic acid- or soil drying-induced stomatal closure in aged wheat leaves.
Plant Cell Environ.
PUBLISHED: 01-06-2013
Show Abstract
Hide Abstract
The mechanism of age-induced decreased stomatal sensitivity to abscisic acid (ABA) and soil drying has been explored here. Older, fully expanded leaves partly lost their ability to close stomata in response to foliar ABA sprays, and soil drying which stimulated endogenous ABA production, while young fully expanded leaves closed their stomata more fully. However, ABA- or soil drying-induced stomatal closure of older leaves was partly restored by pretreating plants with 1-methylcyclopropene (1-MCP), which can antagonize ethylene receptors, or by inoculating soil around the roots with the rhizobacterium Variovorax paradoxus?5C-2, which contains 1-aminocyclopropane-1-carboxylic acid (ACC)-deaminase. ACC (the immediate biosynthetic precursor of ethylene) sprays revealed higher sensitivity of stomata to ethylene in older leaves than younger leaves, despite no differences in endogenous ACC concentrations or ethylene emission. Taken together, these results indicate that the relative insensitivity of stomatal closure to ABA and soil drying in older leaves is likely due to altered stomatal sensitivity to ethylene, rather than ethylene production. To our knowledge, this is the first study to mechanistically explain diminished stomatal responses to soil moisture deficit in older leaves, and the associated reduction in leaf water-use efficiency.
Related JoVE Video
Omics of root-to-shoot signaling under salt stress and water deficit.
OMICS
PUBLISHED: 12-02-2011
Show Abstract
Hide Abstract
Maximizing crop yield depends on the leaves receiving an optimal supply of water, mineral nutrients, small organic molecules, proteins, and hormones from the root system via the xylem. Soil drying and salinization alter these xylem fluxes, and modern omics techniques offer unparalleled opportunities to understand the complexity of these responses. Although absolute xylem concentrations of any constituent depend on the genotype and xylem sap sampling methodology, analysis of the relative changes in concentrations has revealed some conserved behavior. Typically, these stresses increase xylem concentrations of the plant hormone abscisic acid (ABA) that limits crop water loss, but decrease the concentrations of certain cytokinins that stimulate expansive growth and prevent premature leaf senescence. Further understanding of the ionic and biophysical alterations in the rhizosphere environment that cause increased xylem concentrations of the ethylene precursor (ACC) is needed. Interactions of these plant hormones with plant nutrient status and xylem nutrient delivery may be important in tuning plant responses to their environment. Xylem proteomics is an emerging area that will help understand mechanisms of plant stress adaptation. Using omics techniques to underpin rootstock-mediate plant improvement is likely to improve crop yields in dry or saline soil.
Related JoVE Video
Genetic and management approaches to boost UK wheat yields by ameliorating water deficits.
J. Exp. Bot.
PUBLISHED: 09-02-2011
Show Abstract
Hide Abstract
Faced with the challenge of increasing global food production, there is the need to exploit all approaches to increasing crop yields. A major obstacle to boosting yields of wheat (an important staple in many parts of the world) is the availability and efficient use of water, since there is increasing stress on water resources used for agriculture globally, and also in parts of the UK. Improved soil and crop management and the development of new genotypes may increase wheat yields when water is limiting. Technical and scientific issues concerning management options such as irrigation and the use of growth-promoting rhizobacteria are explored, since these may allow the more efficient use of irrigation. Fundamental understanding of how crops sense and respond to multiple abiotic stresses can help improve the effective use of irrigation water. Experiments are needed to test the hypothesis that modifying wheat root system architecture (by increasing root proliferation deep in the soil profile) will allow greater soil water extraction thereby benefiting productivity and yield stability. Furthermore, better knowledge of plant and soil interactions and how below-ground and above-ground processes communicate within the plant can help identify traits and ultimately genes (or alleles) that will define genotypes that yield better under dry conditions. Developing new genotypes will take time and, therefore, these challenges need to be addressed now.
Related JoVE Video
Microbial enhancement of crop resource use efficiency.
Curr. Opin. Biotechnol.
PUBLISHED: 08-01-2011
Show Abstract
Hide Abstract
Naturally occurring soil microbes may be used as inoculants to maintain crop yields despite decreased resource (water and nutrient) inputs. Plant symbiotic relationships with mycorrhizal fungi alter root aquaporin gene expression and greatly increase the surface area over which plant root systems take up water and nutrients. Soil bacteria on the root surface alter root phytohormone status thereby increasing growth, and can make nutrients more available to the plant. Combining different classes of soil organism within one inoculant can potentially take advantage of multiple plant growth-promoting mechanisms, but biological interactions between inoculant constituents and the plant are difficult to predict. Whether the yield benefits of such inocula allow modified nutrient and water management continues to challenge crop biotechnologists.
Related JoVE Video
Barriers and silencers: a theoretical toolkit for control and containment of nucleosome-based epigenetic states.
J. Mol. Biol.
PUBLISHED: 06-16-2011
Show Abstract
Hide Abstract
Positive feedback in nucleosome modification has been proposed to allow large chromatin regions to exist stably and heritably in distinct expression states. However, modeling has shown that such epigenetic bistability requires that modifying enzymes recruited by nucleosomes are active on distant nucleosomes, potentially allowing uncontrollable spreading of modification. By modeling the silencing of mating-type loci in Saccharomyces cerevisiae, we show that a modification reaction that combines a long-range component and a locally acting component can provide bistability and can be blocked by simple barriers that interrupt the nucleosome chain. We find that robust containment of the silenced region could be achieved by the presence of a number of weak simple barriers in the surrounding chromatin and a limited capacity of the positive feedback reaction. In addition, we show that the state of the silenced region can be regulated by silencer elements acting only on neighboring nucleosomes. Thus, a relatively simple set of nucleosome-modifying enzymes and recognition domains is all that is needed to make chromatin-based epigenetics useful and safe.
Related JoVE Video
Root-to-shoot signalling when soil moisture is heterogeneous: increasing the proportion of root biomass in drying soil inhibits leaf growth and increases leaf abscisic acid concentration.
Plant Cell Environ.
PUBLISHED: 04-21-2011
Show Abstract
Hide Abstract
To determine whether root-to-shoot signalling of soil moisture heterogeneity depended on root distribution, wild-type (WT) and abscisic acid (ABA)-deficient (Az34) barley (Hordeum vulgare) plants were grown in split pots into which different numbers of seminal roots were inserted. After establishment, all plants received the same irrigation volumes, with one pot watered (w) and the other allowed to dry the soil (d), imposing three treatments (1 d: 3 w, 2 d: 2 w, 3 d: 1 w) that differed in the number of seminal roots exposed to drying soil. Root distribution did not affect leaf water relations and had no sustained effect on plant evapotranspiration (ET). In both genotypes, leaf elongation was less and leaf ABA concentrations were higher in plants with more roots in drying soil, with leaf ABA concentrations and water potentials 30% and 0.2 MPa higher, respectively, in WT plants. Whole-pot soil drying increased xylem ABA concentrations, but maximum values obtained when leaf growth had virtually ceased (100 nm in Az34, 330 nm in WT) had minimal effects (<40% leaf growth inhibition) when xylem supplied to detached shoots. Although ABA may not regulate leaf growth in vivo, genetic variation in foliar ABA concentration in the field may indicate different root distributions between upper (drier) and lower (wetter) soil layers.
Related JoVE Video
Identification of residues in the N-terminal PAS domains important for dimerization of Arnt and AhR.
Nucleic Acids Res.
PUBLISHED: 01-17-2011
Show Abstract
Hide Abstract
The basic helix-loop-helix (bHLH).PAS dimeric transcription factors have crucial roles in development, stress response, oxygen homeostasis and neurogenesis. Their target gene specificity depends in part on partner protein choices, where dimerization with common partner Aryl hydrocarbon receptor nuclear translocator (Arnt) is an essential step towards forming active, DNA binding complexes. Using a new bacterial two-hybrid system that selects for loss of protein interactions, we have identified 22 amino acids in the N-terminal PAS domain of Arnt that are involved in heterodimerization with aryl hydrocarbon receptor (AhR). Of these, Arnt E163 and Arnt S190 were selective for the AhR/Arnt interaction, since mutations at these positions had little effect on Arnt dimerization with other bHLH.PAS partners, while substitution of Arnt D217 affected the interaction with both AhR and hypoxia inducible factor-1? but not with single minded 1 and 2 or neuronal PAS4. Arnt uses the same face of the N-terminal PAS domain for homo- and heterodimerization and mutational analysis of AhR demonstrated that the equivalent region is used by AhR when dimerizing with Arnt. These interfaces differ from the PAS ?-scaffold surfaces used for dimerization between the C-terminal PAS domains of hypoxia inducible factor-2? and Arnt, commonly used for PAS domain interactions.
Related JoVE Video
Root-targeted biotechnology to mediate hormonal signalling and improve crop stress tolerance.
Plant Cell Rep.
PUBLISHED: 01-07-2011
Show Abstract
Hide Abstract
Since plant root systems capture both water and nutrients essential for the formation of crop yield, there has been renewed biotechnological focus on root system improvement. Although water and nutrient uptake can be facilitated by membrane proteins known as aquaporins and nutrient transporters, respectively, there is a little evidence that root-localised overexpression of these proteins improves plant growth or stress tolerance. Recent work suggests that the major classes of phytohormones are involved not only in regulating aquaporin and nutrient transporter expression and activity, but also in sculpting root system architecture. Root-specific expression of plant and bacterial phytohormone-related genes, using either root-specific or root-inducible promoters or grafting non-transformed plants onto constitutive hormone producing rootstocks, has examined the role of root hormone production in mediating crop stress tolerance. Root-specific traits such as root system architecture, sensing of edaphic stress and root-to-shoot communication can be exploited to improve resource (water and nutrients) capture and plant development under resource-limited conditions. Thus, root system engineering provides new opportunities to maintain sustainable crop production under changing environmental conditions.
Related JoVE Video
Root-synthesized cytokinins improve shoot growth and fruit yield in salinized tomato (Solanum lycopersicum L.) plants.
J. Exp. Bot.
PUBLISHED: 10-19-2010
Show Abstract
Hide Abstract
Salinity limits crop productivity, in part by decreasing shoot concentrations of the growth-promoting and senescence-delaying hormones cytokinins. Since constitutive cytokinin overproduction may have pleiotropic effects on plant development, two approaches assessed whether specific root-localized transgenic IPT (a key enzyme for cytokinin biosynthesis) gene expression could substantially improve tomato plant growth and yield under salinity: transient root IPT induction (HSP70::IPT) and grafting wild-type (WT) shoots onto a constitutive IPT-expressing rootstock (WT/35S::IPT). Transient root IPT induction increased root, xylem sap, and leaf bioactive cytokinin concentrations 2- to 3-fold without shoot IPT gene expression. Although IPT induction reduced root biomass (by 15%) in control (non-salinized) plants, in salinized plants (100?mM NaCl for 22?d), increased cytokinin concentrations delayed stomatal closure and leaf senescence and almost doubled shoot growth (compared with WT plants), with concomitant increases in the essential nutrient K(+) (20%) and decreases in the toxic ion Na(+) (by 30%) and abscisic acid (by 20-40%) concentrations in transpiring mature leaves. Similarly, WT/35S::IPT plants (scion/rootstock) grown with 75?mM NaCl for 90?d had higher fruit trans-zeatin concentrations (1.5- to 2-fold) and yielded 30% more than WT/non-transformed plants. Enhancing root cytokinin synthesis modified both shoot hormonal and ionic status, thus ameliorating salinity-induced decreases in growth and yield.
Related JoVE Video
Economy of operon formation: cotranscription minimizes shortfall in protein complexes.
MBio
PUBLISHED: 09-21-2010
Show Abstract
Hide Abstract
Genes of prokaryotes and Archaea are often organized in cotranscribed groups, or operons. In contrast, eukaryotic genes are generally transcribed independently. Here we show that there is a substantial economic gain for the cell to cotranscribe genes encoding protein complexes because it synchronizes the fluctuations, or noise, in the levels of the different components. This correlation substantially reduces the shortfall in production of the complex. This benefit is relatively large in small cells such as bacterial cells, in which there are few mRNAs and proteins per cell, and is diminished in larger cells such as eukaryotic cells.
Related JoVE Video
Minimal gene regulatory circuits for a lysis-lysogeny choice in the presence of noise.
PLoS ONE
PUBLISHED: 08-04-2010
Show Abstract
Hide Abstract
Gene regulatory networks (GRNs) that make reliable decisions should have design features to cope with random fluctuations in the levels or activities of biological molecules. The phage ? GRN makes a lysis-lysogeny decision informed by the number of phages infecting the cell. To analyse the design of decision making GRNs, we generated random in silico GRNs comprised of two or three transcriptional regulators and selected those able to perform a ?-like decision in the presence of noise. Various two-protein networks analogous to the ? CI-Cro GRN worked in noise-less conditions but failed when noise was introduced. Adding a ? CII-like protein significantly improved robustness to noise. CII relieves the CI-like protein of its decider function, allowing CI to be optimized as a decision maintainer. CIIs lysogenic decider function was improved by its instability and rapid removal once the decision was taken, preventing its interference with maintenance. A more reliable decision also resulted from simulated co-transcription of the genes for CII and the Cro-like protein, which correlates fluctuations in these opposing decider functions and makes their ratio less noisy. Thus, the ? decision network contains design features for reducing and resisting noise.
Related JoVE Video
Principal component analysis of hormone profiling data suggests an important role for cytokinins in regulating leaf growth and senescence of salinized tomato.
Plant Signal Behav
PUBLISHED: 07-02-2010
Show Abstract
Hide Abstract
High throughput analytical methods allow phytohormonal profiling, but the magnitude of the data generated makes it difficult to draw firm conclusions about the physiological roles of different compounds. Principal component analysis (PCA) was used as a mathematical tool to evaluate relationships between physiological and hormonal variables in two experiments with salinised tomato. When tomato plants (cv Boludo F1) were grafted onto a recombinant inbred line (RIL) population derived from a Solanum lycopersicum x S. cheesmaniae cross and grown under moderate salinity (75 mM NaCl) for 100 days under greenhouse conditions, PCA revealed an important role for leaf xylem cytokinins (CKs) in controlling leaf growth and photosystem II efficiency (Fv/Fm) and thus crop productivity under salinity. PCA analysis from a similar experiment, with ungrafted tomato grown under highly saline (100 mM NaCl) conditions, that evaluated the temporal sequence of leaf growth (as relative growth rate, LRGR) and senescence and hormone concentrations, revealed a similar influence of CKs on both processes, since Fv/Fm and LRGR were strongly loaded along the two principal components and placed in the same cluster as leaf trans-zeatin and/or related to other CK-related parameters. The conservative behaviour of the eigen vectors for Fv/Fm and the analyzed phytohormones in different compartments (xylem, leaf and root) between different experiments suggests an important role for CKs in regulating leaf senescence, while CKs and other hormones seem to regulate leaf growth under salinity.
Related JoVE Video
Root water potential integrates discrete soil physical properties to influence ABA signalling during partial rootzone drying.
J. Exp. Bot.
PUBLISHED: 06-30-2010
Show Abstract
Hide Abstract
To investigate the influence of different growing substrates (two mineral, two organic) on root xylem ABA concentration ([ABA](root)) and the contribution of the drying root system to total sap flow during partial rootzone drying (PRD), sunflower (Helianthus annuus L.) shoots were grafted onto the root systems of two plants grown in separate pots. Sap flow through each hypocotyl was measured below the graft union when one pot (wet) was watered and other (dry) was not. Each substrate gave unique relationships between dry pot matric potential (Psi(soil)), volumetric water content ((v)) or penetrometer resistance (Q) and either the fraction of photoperiod sap flow from roots in drying soil or [ABA](root). However, decreased relative sap flow, and increased [ABA](root), from roots in drying soil varied with root water potential (Psi(root)) more similarly across a range of substrates. The gradient between Psi(soil) and Psi(root) was greater in substrates with high sand or peat proportions, which may have contributed to a more sensitive response of [ABA](root) to Psi(soil) in these substrates. Whole plant transpiration was most closely correlated with the mean Psi(soil) of both pots, and then with detached leaf xylem ABA concentration. Although Psi(root) best predicted decreased relative sap flow, and increased [ABA](root), from roots in drying soil across a range of substrates, the inaccessibility of this variable in field studies requires a better understanding of how measurable soil variables (Psi(soil), (v), Q) affect Psi(root).
Related JoVE Video
Theory for the stability and regulation of epigenetic landscapes.
Phys Biol
PUBLISHED: 06-08-2010
Show Abstract
Hide Abstract
Cells can often choose among several stably heritable phenotypes. Examples are the expressions of genes in eukaryotic cells where long chromosomal regions can adopt persistent and heritable silenced or active states that may be associated with positive feedback in dynamic modification of nucleosomes. We generalize this mechanism in terms of bistability associated with valleys in an epigenetic landscape. A transfer matrix method was used to rigorously follow the system through the disruptive process of cell division. This combined treatment of noisy dynamics both between and during cell division provides an efficient way to calculate the stability of alternative states in a broad range of epigenetic systems.
Related JoVE Video
Genetic analysis of physiological components of salt tolerance conferred by Solanum rootstocks. What is the rootstock doing for the scion?
Theor. Appl. Genet.
PUBLISHED: 02-01-2010
Show Abstract
Hide Abstract
Grafting desirable crop varieties on stress-tolerant rootstocks provides an opportunity to increase crop salt tolerance. Here, a commercial hybrid tomato variety was grafted on two populations of recombinant inbred lines developed from a salt-sensitive genotype of Solanum lycopersicum var. cerasiforme, as female parent, and two salt-tolerant lines, as male parents, from S. pimpinellifolium, the P population, and S. cheesmaniae, the C population, to identify an easy screening method for identifying rootstocks conferring salt tolerance in terms of fruit yield. Potential physiological components of salt tolerance were assessed in the scion: leaf biomass, [Na(+)], nutrition, water relations and xylem ABA concentration. A significant correlation between scion fruit yield and scion leaf fresh weight, water potential or the ABA concentration was found in the C population under salinity, but the only detected QTL did not support this relationship. The rootstocks of the P population clearly affected seven traits related to the sodium, phosphorous and copper concentrations and water content of the scion leaf, showing heritability estimates around 0.4 or higher. According to heritability estimates in the P population, up to five QTLs were detected per trait. QTLs contributing over 15% to the total variance were found for P and Cu concentrations and water content of the scion leaf, and the proportion of fresh root weight. Correlation and QTL analysis suggests that rootstock-mediated improvement of fruit yield in the P population under salinity is mainly explained by the rootstocks ability to minimise perturbations in scion water status.
Related JoVE Video
Why do phage play dice?
J. Virol.
PUBLISHED: 09-09-2009
Show Abstract
Hide Abstract
Phage lambda is among the simplest organisms that make a developmental decision. An infected bacterium goes either into the lytic state, where the phage particles rapidly replicate and eventually lyse the cell, or into a lysogenic state, where the phage goes dormant and replicates along with the cell. Experimental observations by P. Kourilsky are consistent with a single phage infection deterministically choosing lysis and double infection resulting in a stochastic choice. We argue that the phage are playing a "game" of minimizing the chance of extinction and that the shift from determinism to stochasticity is due to a shift from a single-player to a multiplayer game. Crucial to the argument is the clonal identity of the phage.
Related JoVE Video
Partial phenotypic reversion of ABA-deficient flacca tomato (Solanum lycopersicum) scions by a wild-type rootstock: normalizing shoot ethylene relations promotes leaf area but does not diminish whole plant transpiration rate.
J. Exp. Bot.
PUBLISHED: 07-31-2009
Show Abstract
Hide Abstract
To evaluate the role of root-synthesized ABA in regulating growth and stomatal behaviour under well-watered conditions, isogenic wild-type (WT) and ABA-deficient flacca (flc) tomato (Solanum lycopersicum) were reciprocally and self-grafted just below the cotyledonary node. Since flc scions had lower leaf water potentials due to higher transpiration rates, a subset of all graft combinations was grown under a shoot misting treatment to minimize differences in shoot water status. Misting did not alter the relative effects of the different graft combinations on leaf area. WT scions had the greatest leaf area and lowest whole plant transpiration rate irrespective of the rootstock, implying that shoot ABA biosynthesis was sufficient to account for a WT shoot phenotype. In WT scions, the rootstock had no effect on detached leaf ethylene evolution or xylem concentrations of ABA or the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC). In flc scions, although the WT rootstock suppressed stomatal conductance of individual leaves, there was no detectable effect on whole plant transpiration rate. However, leaf area of flc/WT (scion/rootstock) plants increased 1.6-fold compared to flc self-grafts. WT rootstocks increased xylem ABA concentration in flc scions (relative to flc self-grafts) up to 3-fold, and resulted in xylem ACC concentrations and detached leaf ethylene evolution similar to WT scions. Since the WT rootstock normalized shoot ethylene relations but only partially restored the leaf area of flc scions (relative to that of WT scions), shoot ABA biosynthesis can directly promote leaf area via an unknown, ethylene-independent, mechanism.
Related JoVE Video
Minimal gene regulatory circuits that can count like bacteriophage lambda.
J. Mol. Biol.
PUBLISHED: 06-17-2009
Show Abstract
Hide Abstract
The behavior of living systems is dependent on large dynamical gene regulatory networks (GRNs). However, the functioning of even the smallest GRNs is difficult to predict. The bistable GRN of bacteriophage lambda is able to count to make a decision between lysis and lysogeny on the basis of the number of phages infecting the cell, even though replication of the phage genome eliminates this initial difference. By simulating the behavior of a large number of random transcriptional GRNs, we show that a surprising variety of GRNs can carry out this complex task, including simple CI-Cro-like mutual repression networks. Thus, our study extends the repertoire of simple GRNs. Counterintuitively, the major effect of the addition of CII-like regulation, generally thought to be needed for counting by lambda, was to improve the ability of the networks to complete a simulated prophage induction. Our study suggests that additional regulatory mechanisms to decouple Cro and CII levels may exist in lambda and that infection counting could be widespread among temperate bacteriophages, many of which contain CI-Cro-like circuits.
Related JoVE Video
Water relations of the tos1 tomato mutant at contrasting evaporative demand.
Physiol Plant
PUBLISHED: 06-09-2009
Show Abstract
Hide Abstract
The tos1 (tomato osmotically sensitive) mutant, isolated from an in vitro screen of root growth during osmotic stress, was less sensitive to exogenous ABA, but accumulated more ABA under osmotic stress than WT plants. We assessed growth and water relations characteristics of hydroponically grown tos1 seedlings (in the absence of osmotic stress) at low and high evaporative demands. Growth of tos1 was severely inhibited at both high and low evaporative demands. Twenty DAS, WT and tos1 genotypes had a similar leaf water and turgor potential, but mature tos1 plants (45 day old) showed a significant diurnal loss of leaf turgor, with recovery overnight. Increased evaporative demand increased turgor loss of tos1 plants. High evaporative demand at the beginning of the day decreased stomatal conductance of tos1, without diurnal recovery, thus whole plant transpiration was decreased. De-topped tos1 seedlings showed decreased root hydraulic conductance and had a 1.4-fold increase in root ABA concentration. Impaired root function of tos1 plants failed to meet transpirational water demand and resulted in shoot turgor loss, stomatal closure and growth inhibition.
Related JoVE Video
Rootstock-mediated changes in xylem ionic and hormonal status are correlated with delayed leaf senescence, and increased leaf area and crop productivity in salinized tomato.
Plant Cell Environ.
PUBLISHED: 03-03-2009
Show Abstract
Hide Abstract
Tomato crop productivity under salinity can be improved by grafting cultivars onto salt-tolerant wild relatives, thus mediating the supply of root-derived ionic and hormonal factors that regulate leaf area and senescence. A tomato cultivar was grafted onto rootstocks from a population of recombinant inbred lines (RILs) derived from a Solanum lycopersicum x Solanum cheesmaniae cross and cultivated under moderate salinity (75 mM NaCl). Concentrations of Na(+), K(+) and several phytohormones [abscisic acid (ABA); the cytokinins (CKs) zeatin, Z; zeatin riboside, ZR; and the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC)] were analysed in leaf xylem sap in graft combinations of contrasting vigour. Scion leaf area correlated with photosystem II (PSII) efficiency (F(v)/F(m)) and determined fruit productivity. Xylem K(+) (but not Na(+)), K(+)/Na(+), the active CK Z, the ratio with its storage form Z/ZR and especially the ratio between CKs and ACC (Z/ACC and Z + ZR/ACC) were positively loaded into the first principal component (PC) determining both leaf growth and PSII efficiency. In contrast, the ratio ACC/ABA was negatively correlated with leaf biomass. Although the underlying physiological mechanisms by which rootstocks mediate leaf area or chlorophyll fluorescence (and thus influence tomato salt tolerance) seem complex, a putative potassium-CK interaction involved in regulating both processes merits further attention.
Related JoVE Video
Potent transcriptional interference by pausing of RNA polymerases over a downstream promoter.
Mol. Cell
PUBLISHED: 02-24-2009
Show Abstract
Hide Abstract
Elongating RNA polymerases (RNAPs) can interfere with transcription from downstream promoters by inhibiting DNA binding by RNAP and activators. However, combining quantitative measurement with mathematical modeling, we show that simple RNAP elongation cannot produce the strong asymmetric interference observed between a natural face-to-face promoter pair in bacteriophage lambda. Pausing of elongating polymerases over the RNAP-binding site of the downstream promoter is demonstrated in vivo and is shown by modeling to account for the increased interference. The model successfully predicts the effects on interference of treatments increasing or reducing pausing. Gene regulation by pausing-enhanced occlusion provides a general and potentially widespread mechanism by which even weak converging or tandem transcription, either coding or noncoding, can bring about strong in cis repression.
Related JoVE Video
Rhizosphere bacteria containing 1-aminocyclopropane-1-carboxylate deaminase increase yield of plants grown in drying soil via both local and systemic hormone signalling.
New Phytol.
PUBLISHED: 01-06-2009
Show Abstract
Hide Abstract
Decreased soil water availability can stimulate production of the plant hormone ethylene and inhibit plant growth. Strategies aimed at decreasing stress ethylene evolution might attenuate its negative effects. An environmentally benign (nonchemical) method of modifying crop ethylene relations - soil inoculation with a natural root-associated bacterium Variovorax paradoxus 5C-2 (containing the enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase that degrades the ethylene precursor ACC), was assessed with pea (Pisum sativum) plants grown in drying soil. Inoculation with V. paradoxus 5C-2, but not with a transposome mutant with massively decreased ACC deaminase activity, improved growth, yield and water-use efficiency of droughted peas. Systemic effects of V. paradoxus 5C-2 included an amplified soil drying-induced increase of xylem abscisic acid (ABA) concentration, but an attenuated soil drying-induced increase of xylem ACC concentration. A local bacterial effect was increased nodulation by symbiotic nitrogen-fixing bacteria, which prevented a drought-induced decrease in nodulation and seed nitrogen content. Successfully deploying a single bacterial gene in the rhizosphere increased yield and nutritive value of plants grown in drying soil, via both local and systemic hormone signalling. Such bacteria may provide an easily realized, economic means of sustaining crop yields and using irrigation water more efficiently in dryland agriculture.
Related JoVE Video
Multiple impacts of the plant growth-promoting rhizobacterium Variovorax paradoxus 5C-2 on nutrient and ABA relations of Pisum sativum.
J. Exp. Bot.
Show Abstract
Hide Abstract
Resolving the physiological mechanisms by which rhizobacteria enhance plant growth is difficult, since many such bacteria contain multiple plant growth-promoting properties. To understand further how the 1-aminocyclopropane-1-carboxylate (ACC) deaminase (ACCd)-containing rhizobacterium Variovorax paradoxus 5C-2 affects plant growth, the flows and partitioning of mineral nutrients and abscisic acid (ABA) and ABA metabolism were studied in pea (Pisum sativum) plants following rhizosphere bacterial inoculation. Although root architecture was not affected, inoculation increased root and shoot biomass, and stomatal conductance, by 20, 15, and 24%, respectively, and increased N, P, K, Ca, and Mg uptake by 16, 81, 50, 46, and 58%, respectively. P deposition in inoculated plant roots was 4.9 times higher than that in uninoculated controls. Rhizobacterial inoculation increased root to shoot xylem flows and shoot to root phloem flows of K by 1.8- and 2.1-fold, respectively. In control plants, major sinks for K deposition were the roots and upper shoot (43% and 49% of total uptake, respectively), while rhizobacterial inoculation increased K distribution to the lower shoot at the expense of other compartments (xylem, phloem, and upper shoot). Despite being unable to metabolize ABA in vitro, V. paradoxus 5C-2 decreased root ABA concentrations and accumulation by 40-60%. Although inoculation decreased xylem ABA flows, phloem ABA flows increased. Whether bacterial ACCd attenuates root to shoot ABA signalling requires further investigation, since ABA is critical to maintain growth of droughted plants, and ACCd-containing organisms have been advocated as a means of minimizing growth inhibition of plants in drying soil.
Related JoVE Video
A simple histone code opens many paths to epigenetics.
PLoS Comput. Biol.
Show Abstract
Hide Abstract
Nucleosomes can be covalently modified by addition of various chemical groups on several of their exposed histone amino acids. These modifications are added and removed by enzymes (writers) and can be recognized by nucleosome-binding proteins (readers). Linking a reader domain and a writer domain that recognize and create the same modification state should allow nucleosomes in a particular modification state to recruit enzymes that create that modification state on nearby nucleosomes. This positive feedback has the potential to provide the alternative stable and heritable states required for epigenetic memory. However, analysis of simple histone codes involving interconversions between only two or three types of modified nucleosomes has revealed only a few circuit designs that allow heritable bistability. Here we show by computer simulations that a histone code involving alternative modifications at two histone positions, producing four modification states, combined with reader-writer proteins able to distinguish these states, allows for hundreds of different circuits capable of heritable bistability. These expanded possibilities result from multiple ways of generating two-step cooperativity in the positive feedback--through alternative pathways and an additional, novel cooperativity motif. Our analysis reveals other properties of such epigenetic circuits. They are most robust when the dominant nucleosome types are different at both modification positions and are not the type inserted after DNA replication. The dominant nucleosome types often recruit enzymes that create their own type or destroy the opposing type, but never catalyze their own destruction. The circuits appear to be evolutionary accessible; most circuits can be changed stepwise into almost any other circuit without losing heritable bistability. Thus, our analysis indicates that systems that utilize an expanded histone code have huge potential for generating stable and heritable nucleosome modification states and identifies the critical features of such systems.
Related JoVE Video
Contrasting physiological effects of partial root zone drying in field-grown grapevine (Vitis vinifera L. cv. Monastrell) according to total soil water availability.
J. Exp. Bot.
Show Abstract
Hide Abstract
Different spatial distributions of soil moisture were imposed on field-grown grapevines by applying the same irrigation volumes to the entire (DI; deficit irrigation) or part of the (PRD; partial root zone drying) root zone. Five treatments were applied: controls irrigated at 60% ETc (crop evapotranspiration) for the whole season (308 mm year(-1)); DI-1 and PRD-1 that received the same irrigation as controls before fruit set, 30% ETc from fruit set to harvest and 45% ETc post-harvest (192 mm year(-1)); and DI-2 and PRD-2 that were the same, except that 15% ETc was applied from fruit set to harvest (142 mm year(-1)). Compared with DI-1, PRD-1 maintained higher leaf area post-veraison and increased root water uptake, whole-plant hydraulic conductance, leaf transpiration, stomatal conductance, and photosynthesis, but decreased intrinsic gas exchange efficiency without causing differences in leaf xylem abscisic acid (ABA) concentration. Compared with DI-2, PRD-2 increased leaf xylem ABA concentration and decreased root water uptake, whole-plant hydraulic conductance, leaf transpiration, stomatal conductance, and photosynthesis, mainly at the beginning of PRD cycles. Distinctive PRD effects (e.g. greater stomatal closure) depended on the volumetric soil water content of the wet root zone, as predicted from a model of root-to-shoot ABA signalling.
Related JoVE Video
Xylem sap collection and extraction methodologies to determine in vivo concentrations of ABA and its bound forms by gas chromatography-mass spectrometry (GC-MS).
Plant Methods
Show Abstract
Hide Abstract
Accurate quantification of xylem sap ABA concentrations is important to underpin models of root-to-shoot ABA signalling to predict the physiological effects of soil drying. Growing tomato plants in a whole plant pressure chamber allowed sequential xylem sap collection from a detached leaf, the petiole stub of an otherwise intact plant and finally the de-topped root system of the same plant, to determine the impact of xylem sap sampling methodology on xylem ABA concentration. Since xylem sap can contain bound forms of ABA, a novel gas chromatography-mass spectrometry (GC-MS) procedure was developed to chemically separate free ABA from two in planta bound ABA forms known as Adducts I and II and ABA-glucose-ester (ABA-GE).
Related JoVE Video
Microbial amelioration of crop salinity stress.
J. Exp. Bot.
Show Abstract
Hide Abstract
The use of soil and irrigation water with a high content of soluble salts is a major limiting factor for crop productivity in the semi-arid areas of the world. While important physiological insights about the mechanisms of salt tolerance in plants have been gained, the transfer of such knowledge into crop improvement has been limited. The identification and exploitation of soil microorganisms (especially rhizosphere bacteria and mycorrhizal fungi) that interact with plants by alleviating stress opens new alternatives for a pyramiding strategy against salinity, as well as new approaches to discover new mechanisms involved in stress tolerance. Although these mechanisms are not always well understood, beneficial physiological effects include improved nutrient and water uptake, growth promotion, and alteration of plant hormonal status and metabolism. This review aims to evaluate the beneficial effects of soil biota on the plant response to saline stress, with special reference to phytohormonal signalling mechanisms that interact with key physiological processes to improve plant tolerance to the osmotic and toxic components of salinity. Improved plant nutrition is a quite general beneficial effect and may contribute to the maintenance of homeostasis of toxic ions under saline stress. Furthermore, alteration of crop hormonal status to decrease evolution of the growth-retarding and senescence-inducing hormone ethylene (or its precursor 1-aminocyclopropane-1-carboxylic acid), or to maintain source-sink relations, photosynthesis, and biomass production and allocation (by altering indole-3-acetic acid and cytokinin biosynthesis) seem to be promising target processes for soil biota-improved crop salt tolerance.
Related JoVE Video

What is Visualize?

JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

How does it work?

We use abstracts found on PubMed and match them to JoVE videos to create a list of 10 to 30 related methods videos.

Video X seems to be unrelated to Abstract Y...

In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.